Heart failure (HF) is a leading cause of morbidity and mortality, contributing to 1 in 9 deaths in the US. Consequently, there is an enormous need for new HF therapies, which can only emerge from discovery of new therapeutic targets. In the past, inotropic drugs that enhance myocardial performance acutely were developed to treat HF, but most of them are now contraindicated because they worsen HF outcomes long-term. Recently, we developed a novel culturing method, termed Matrigel Mattress, which allowed the simultaneous assessment of contractile performance and calcium dynamics in individual human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Using the Matrigel Mattress method as a basis for a chemical screening platform, we discovered that the small molecule EGM significantly enhanced both inotropy and lusitropy in hiPSC-CMs and improved cardiac function in vivo. Unlike the traditional inotropes, EGM did not affect calcium (Ca) cycling, cellular cAMP concentrations or increase the beat rate, suggesting it acts by a fundamentally novel mechanism. To unlock the mechanistic underpinnings of EGM's pharmacology, we carried out a biochemical pull-down assay and identified farnesyl diphosphate synthase (FDPS), required for protein prenylation, as a candidate target of EGM. Consistent with prior studies demonstrating that FDPS contributes to hypertrophy and HF in animal models, we found that naturally occurring variants in the FDPS gene were highly associated with HF in Vanderbilt University Medical Center's electronic health record-linked DNA database. The latter result, based on real world clinical data, raises the exciting possibility that modulating the level of FDPS activity over a course of a person's life can significantly alter HF natural history; and that compounds like EGM that inhibit FDPS may improve long-term HF outcomes. Based on these findings, we hypothesize that EGM enhances myocardial performance by inhibiting FDPS, and that FDPS inhibition improves both acute cardiac function and long-term HF outcome. Here, we propose innovative chemical and functional genomic approaches to elucidate the role of FDPS in EGM function. In Aim 1, we will carry out a structure activity relationship (SAR) study of EGM analogs to determine whether FDPS inhibition is essential for EGM function. In Aim 2, we will employ the CRISPR/Cas9-mediated genome editing to determine whether ablating the FDPS gene recapitulates EGM's unique pharmacology in hiPSC-CMs. In Aim 3, we will utilize the CRISPR/Cas9- directed homology directed repair (HDR) to introduce the nucleotide changes corresponding to the CHF- associated FDPS variants, and evaluate their impact on hiPSC-CM performance and FDPS function. The proposed study will delineate the effects of FDPS modulation on myocardial performance, and possibly identify additional targets of EGM. This study leverages the unique pharmacology of EGM to lay the foundation for a new understanding of myocardial regulation and the new therapeutic paradigm of ?dual purpose? drugs that acutely relieve HF symptoms as well as improve long-term HF outcomes.